Expression of eukaryotic membrane proteins in eukaryotic and prokaryotic hosts

Abstract

The production of membrane proteins of high purity and in satisfactory yields is crucial for biomedical research. Due to their involvement in various cellular processes, membrane proteins have increasingly become some of the most important drug targets in modern times. Therefore, their structural and functional characterization is a high priority. However, protein expression has always been more challenging for membrane proteins than for soluble proteins. In this review, we present four of the most commonly-used expression systems for eukaryotic membrane proteins. We describe the benefits and drawbacks of bacterial, yeast, insect and mammalian cells. In addition, we describe the different features (growth rate, yield, post-translational modifications) of each expression system, and how they are influenced by the construct design and modifications of the target gene. Cost-effective and fast-growing E. coli is mostly selected for the production of small, simple membrane proteins that, if possible, do not require post-translational modifications but has the potential for the production of bigger proteins as well. Yeast hosts are advantageous for larger and more complex proteins but for the most complex ones, insect or mammalian cells are used as they are the only hosts able to perform all the post-translational modifications found in human cells. A combination of rational construct design and host cell choice can dramatically improve membrane protein production processes.

Identification and Functional Analysis of Cytokine-Like Protein CLEC-47 in Caenorhabditis elegans

“…However, many commercially available recombinant cytokines were produced in the prokaryotic system (R&D Systems), and they still have relative activities. Therefore, in the current study, we chose to express recombinant CLEC-47 in a prokaryotic expression system for protein purification that has the advantage of lower cost and higher yield than HEK 293 eukaryotic cell expression systems ( 42 , 43 ).…”

The Hypothetical Inclusion Membrane Protein CPSIT_0846 Regulates Mitochondrial-Mediated Host Cell Apoptosis via the ERK/JNK Signaling Pathway

“…Some domains of these secreted inclusion membrane proteins may expose into the cytoplasm and combine with molecular chaperone proteins to promote folding or delivery, ultimately achieve the purpose of manipulating host cells (Weber and Faris, 2018). The hypothetical inc protein CPSIT_0846 used in this study is a supernatant protein expressed by a prokaryotic expression system, which usually does not interfere with the native conformation of the protein (Kesidis et al, 2020). But the mechanism of how CPSIT_0846 enter host cells and which domains are exposed into the cytoplasm and which molecular chaperones are bound with them are still unclear.…”

Native and non-native host assessment towards metabolic pathway reconstructions of plant natural products

“…Similarly, anthocyanin production was enhanced by regulating the expression of flavonol synthase [ 89 ]. The transformation efficiency is dependent on various factors such as the promoters, driving the transgene expression and sequences ( e.g ., Kozak), regulating the translational initiation [ 90 ]. In some cases, the codon usage is modified in the transgene in order to avoid cryptic introns and other instability issues.…”